Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-12 Thread Ali
Hi David! I played with the fragment mass tolerance but I am still having the issue. However I searched the problematic files with TPP's X!Tandem (instead of GPM's) and it worked perfectly with Peptide Prophet! Is there a linux version of it which I can use on the cluster or is it just usable

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-12 Thread Ali
Thanks David! Let me try lowering fragment mass error. I will let you know about the outcome. Well, we already had the Proteome Discoverer results and we really want to create the spectral library asap, so I was given those results to use instead of X!Tandem. and I have never worked with

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-12 Thread David Shteynberg
The TPP is not tested to work with ProteomeDiscoverer pepXML. Sure you can send me a sample file, but it may take some time to implement support for this type of data. Comet is open source and easy to use out of the box. Why don't you try that route? -David On Wed, Oct 12, 2016 at 3:24 PM, Ali

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-12 Thread Ali
Dear David Thank you very much! You were right, the problem was with X!Tandem. I decided do the searches with Proteome Discoverer. I exported the results to pep.xml (from .msf) using PD but now the results don't work with PeptideProphet. I can open them in PepXMLViewer but as I said in another

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-12 Thread David Shteynberg
My suspicion regarding the Tandem failure is the fragment mass tolerance. I am observing fragment mass differences of much less than 0.05 daltons, yet your Tandem search uses 0.5 daltons as the tolerance. Could GPM-tandem be having a problem there? Can you try to lower the fragment mass

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-12 Thread David Shteynberg
Hi Ali, I searched your data with Comet and PeptideProphet got valid PSMs in the analysis that failed to produce correct results from your GPM-version of tandem on the same data. The problem in your dataset is the Tandem search. Attached are the comet parameters I used to search your data.

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-05 Thread Ali
I sent the link to your email. Thank you very much! On Wednesday, October 5, 2016 at 9:21:04 PM UTC-4, David Shteynberg wrote: > > I cannot tell from the search parameters if there is a problem. The only > issue I saw was that you didn't enable isotopic offsets, which are > sometimes helpful.

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-05 Thread Ali
How can I send you these files? Can I upload them on the cloud and send you the link? I am not doing the mass spec so I don't know the details of each experiments but some of them have all three charges and some of them only have 2+ and 3+. Here is an example of similar issue for a file with

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-05 Thread David Shteynberg
I cannot tell from the search parameters if there is a problem. The only issue I saw was that you didn't enable isotopic offsets, which are sometimes helpful. If you send me a sample mzML file and possibly the search results in pep.xml along with the database I can troubleshoot further. One

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-05 Thread Ali
Hi David We are only collecting 2+, 3+ and 4+ data. I am working on the cluster, so I am using the GPM version and we were unable to install the k-score plugin therefore I am using the X! Tandem native scoring for the searches. If you mean combining all of the search results with xinteract

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-05 Thread David Shteynberg
Hi Ali, Are you setting your instrument to collect only 2+ and 3+ data? Are you using the tandem bundled with TPP or the GPM version of tandem? Have you tried to process all your runs in one file? Thanks, -David On Wed, Oct 5, 2016 at 11:39 AM, Ali wrote: > Thank

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-05 Thread Ali
Thank you very much! I really appreciate it! On Tuesday, October 4, 2016 at 8:03:27 PM UTC-4, David Shteynberg wrote: > > PeptideProphet is doing it's job here by telling you that there is no > distinct population of correct results in this search. I suspect your > search parameters are the

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-04 Thread David Shteynberg
PeptideProphet is doing it's job here by telling you that there is no distinct population of correct results in this search. I suspect your search parameters are the problem. This is orbitrap data so the charges should be known, but it seems Tandem only searched 2+ and 3+ for you, this is

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-04 Thread Ali
It appears that PeptideProphet fails, I am getting this message: using Accurate Mass Bins using PPM mass difference (X! Tandem) (minprob 0) adding Accurate Mass mixture distr init with X! Tandem trypsin MS Instrument info: Manufacturer: Thermo Scientific, Model: UNKNOWN, Ionization:

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-04 Thread Ali
Thanks for your recommendations David. I added decoys to my data base and I am still getting 0 probabilities for all of the hits. I am attaching my X! Tandem search parameters xml, could you please kindly do me a big favor and check it to see if there is anything suspicious there which might

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-03 Thread David Shteynberg
I highly recommend you start employing decoys in your search databases. I noticed you are not combining together the runs in the analysis. If the data was generated and searched in the same way it should be analysed together for better statistical power. I suggest you combine the results from

Re: [spctools-discuss] PeptideProphet returns zero probability for all of the X!Tandem search results

2016-10-03 Thread David Shteynberg
Is it possible there are no correct results in that set? Common pitfalls are incorrect search parameters (e.g. wrong mods enabled or disabled) and wrong database. If you have decoys in your database these can be utilized for a more accurate mixture model generation using options "DECOY=